1. Field of the Invention
The present invention relates to a method for expressing hepatitis B viral antigens and virions in vivo, especially relates to a recombinant plasmid for expressing all of the hepatitis B viral antigens and a method to construct an immunocompetent mouse model for persistently expressing hepatitis B vial antigens by using the recombinant plasmid.
2. The Prior Arts
It is estimated that 350 million people are chronic carriers of hepatitis B virus worldwide, and there are approximately 1 million deaths each year caused by hepatitis B related liver diseases. Evidently, hepatitis B virus has tremendous impact on human health, therefore research for best treatment and new drug discovery enthusiastically become an important healthcare issue.
Hepatitis B virus (HBV) is a partially double-stranded DNA virus that infects human hepatocytes. Upon infection of host individual, the hepatitis B virus will first form a covalently closed circular DNA (cccDNA) within the cell, and then followed by a complicated viral replication process. Host individual infected by hepatitis B virus normally develops acute or chronic necroinflammatory of liver diseases, which consequently may transform into hepatocellular carcinoma. It is generally believed that HBV is not directly cytopathic to the hepatocytes, but HBV mediates the immune response of the infected hepatocytes and triggers the infected cells to express viral antigen, which resulting in host immune system attacking its own infected hepatocytes and eventually leading to cell apotosis.
It is necessary and important to develop an appropriate animal model as a tool in search for new therapy and drug discovery, thus HBV transgenic mice have been proposed to serve as animal models in search for treatment of hepatitis and new drug discovery. The whole-genome HBV transgenic mice are created by delivery of a replication-competent HBV genome into mouse chromosome, in which the inserted viral genome is stably and persistently expressed in the liver tissue. However, it has been found that the mechanism of liver inflammation derived from such HBV transgenic mice is different from scenario of human hepatitis. Furthermore, such HBV transgenic mice are HBV antigen tolerant, thus do not exhibit normal immune response against HBV. Hence, it is concluded that such animal models are not ideal in study of HBV tolerance and clearance.
Chisari et al. devise an approach: by injecting hydrodynamically a recombinant plasmid containing the HBV whole genome into immunologically competent mice, to stimulate HBV replication and to induce hepatitis resembling infection process in human. It is proposed that construction of such readily manipulable animal models may be used for studies of HBV induced immune response and hepatitis. (Chisari et al., (2002) Proc. Natl. Acad. Sci. USA 99 (21), 13825-30). In their study, an inbred mouse B10.D2 that shows strong immune response to the HBV surface antigen is selected as a recipient, and pT-MCS-HBV1.3 plasmid is used as a vector to carry HBV gene. Chisari et al. successfully introduce the recombinant plasmid into mice and elicit immune response that resembles human hepatitis after HBV infection. However, there are disadvantages to this animal model, that is, short-term acute liver inflammatory response and clearance of immune system effect 14 days after transfection. These constraints limit application of such animal model in studies such as long term monitoring of immune response after HBV infection and drug evaluation in chronic hepatitis.